With the goal of overcoming the significant delay in neutrophil recovery that occurs following transplantation with umbilical cord blood (CB), we have successfully developed a novel and clinically feasible methodology utilizing an engineered Notch ligand for the ex vivo generation of increased numbers of CD34+ cells. A Phase I clinical trial utilizing Notch-mediated expanded CB progenitors in patients undergoing a myeloablative cord blood transplant (CBT) is currently underway. Results from this trial have not only demonstrated the safety of this approach, but more importantly have for the first time demonstrated that rapid myeloid engraftment can be achieved following infusion of ex vivo expanded hematopoietic progenitors. A significant reduction in median time to an absolute neutrophil count of 500/<ml to just 16 days has been observed in patients receiving expanded cells as compared to a median time of 26 days (p=0.002) in a concurrent cohort of 20 patients undergoing identical treatment but with two non-manipulated CB units. Thus, although the number of patients treated to date is small (n=10), a significant effect on time to myeloid recovery has been clearly demonstrated, as has the safety and clinical feasibility of this approach. Further advancement of this methodology now depends on determination of clinical efficacy, which will require a randomized multi-institutional trial. Prior to initiation of such a trial, several critical issues will need to be addressed, including the ability to manufacture and safely ship cells to outside centers, establishment of an appropriate clinical trials and cell therapy manufacturing collaborative group(s) to conduct a future Phase II randomized multi-institutional trial, and determination of need for HLA-matching of the ex vivo expanded product for clinical use. It is our goal that at the end of the two year funding period, we will be prepared and completely ready to begin critical phase II studies. Specifically, we now propose the following aims: 1) Determine methods for shipping expanded cells to outside centers for infusion, including preclinical evaluation of in vivo repopulating ability in our established murine model of the proposed methods. Following this, we will conduct a 10 patient pilot study as an extension of our current Phase I trial with Dr. John Wagner at the University of Minnesota to evaluate the safety and feasibility of overnight shipment of cells. 2) In a concurrent pilot study to be conducted locally, determine whether infusion of the expanded cell product, which is devoid of T cells, can be infused without need for HLA- matching. In this study, patients undergoing conventional CBT will receive CB progenitors previously expanded from a fresh CB unit and then cryopreserved for future use as an "off-the-shelf" product. If successful, this would dramatically increase patient access to this product. 3) Establish a clinical trials group and cell therapy manufacturing collaborations required to conduct a randomized, multi-institutional trial in two years time via the BMT-CTN and PACT if possible. PUBLIC HEALTH RELEVANCE: Patients undergoing a cord blood transplant, who are often of minority or mixed ethnicity background, are at increased risk of infection and early death following the transplant due to the significant delay in white blood cells recovery (in particular, neutrophils) that these patients experience. However, using a novel culture methodology, we have demonstrated for the first time the ability to generate increased numbers of cells from a single unit of cord blood that are capable of rapid neutrophil recovery when infused in the clinical setting. Further development of this product to confirm our initial promising results requires additional clinical trials that, if successful, could change the way cord blood transplantation is performed.